Normally, when a person's immune system encounters a protein made by a virus or other microbe, fewer than one in 10,000 of the white blood cells known as T lymphocytes react. Although their number is small, these T lymphocytes orchestrate an attack that specifically targets the alien protein, or antigen, without harming healthy tissue. In contrast, proteins called superantigens highly activate the immune system and can cause an unproductive, even destructive, immune response.
Superantigens are the most powerful T cell mitogens known (Johnson, H. M., H. I. Magazine [1988] Int. Arch Allergy Appl. Immunol. 87:87-90). As explained below, these unique antigens stimulate T cells by first binding to class II major histocompatibility (MHC) molecules (Carlsson, R., H. Fischer, H. O. Sjogren [1988] J. Immunol. 140:2484-2488; Fleischer, B., H. Schrezenmeier [1988] J. Exp. Med. 167:1697-1707; Mollick, J. A., R. G. Cook, R. R. Rich [1989] Science 244:817-820) and then as a binary complex bind in a V.sub..beta. specific manner to the T cell antigen receptor (TCR) (Janeway, C. A., J. Yagi, P. J. Conrad, M. E. Katz, B. Jones, S. Vroegop, S. Buxser [1989] Immunol. Rev. 107:61-88; White, J., A. Herman, A. M. Pullen, R. Kubo, J. W. Kappler, P. Marrack [1989] Cell 56:27-35).
Superantigens can arouse as many as one in five T cells, most of which are useless for fighting a current infection. What is worse, certain of the activated cells may unleash an autoimmune attack which targets tissues of the host organism. At times, superantigens may even have the opposite effect: they somehow trigger the death of the cells they excite.
Scientists have gleaned much of what they understand about superantigens from studying the earliest known examples: a group of structurally related proteins called staphylococcal enterotoxins (SEs). Staphylococcal enterotoxins account for as much as 45 percent of all cases of food poisoning. It is well established that when strains of the bacterium Staphylococcus aureus colonize food, they secrete one or more enterotoxins, which are now named alphabetically as A, B, C, D, and E. Within hours after people ingest badly contaminated, toxin-laden food, they begin to feel weak, feverish, and nauseated. Interestingly, intestinal tissue of affected patients looks virtually normal under the microscope. The only obvious abnormality is the presence of white cells in the tissue. It has now also been found that introduction of an enterotoxin to blood triggers the proliferation of lymphocytes. Just a few hundred molecules of toxin triggers a degree of replication that surpasses what could be achieved by a billion copies of a conventional antigen--for example, a protein on the influenza virus.
Further research has documented that a small amount of an enterotoxin can yield extraordinarily high production of chemical signals known as cytokines, which are produced by the subset of T lymphocytes called helper cells. These cells direct much of the immune response. The helper cells do not attack microbes themselves; instead they rely on the cytokines to activate both cytotoxic T lymphocytes, which kill infected cells, and B lymphocytes, which secrete antibodies against antigens.
By the mid-1980s it was recognized that when a tiny amount of enterotoxin A was mixed with T lymphocytes, the collection of cells produced a huge quantity of the cytokine known as interleukin-2 (IL-2). It has also been determined that infusion of high doses of IL-2 into the circulation of cancer patients (as part of an experimental therapy) causes the very symptoms associated with food poisoning. These data indicate that enterotoxins make people ill by stimulating production of high levels of interleukin-2.
Before helper T cells can recognize conventional protein antigens, the proteins must first undergo processing by macrophages or other antigen-presenting cells. These cells engulf antigens and process them into peptides. The presenters then display the peptide antigens at the cell surface in combination with major histocompatibility complex (MHC) class II molecules. A peptide fits in a cleft on an MHC molecule. Once an antigen is displayed, the few helper cells in the body that bear receptors for the particular peptide link up with it. Each T cell is specific for one kind of peptide antigen.
Like conventional toxins, enterotoxin superantigens can arouse helper cells only if antigen-presenting cells display the proteins to the T cells. Moreover, it is MHC class II molecules that do the presenting. Yet, unlike typical antigens, the enterotoxins bind MHC molecules directly; they do not require uptake and processing. Also, enterotoxins do not bind to the inner surface of the peptide-recognizing pocket of the MHC molecule, attaching instead to its outer surface. Then the MHC-superantigen unit contacts the T cell receptor at a site distinct from the surface that envelops conventional antigens. To be more precise, T cell receptors consist of two protein chains, alpha and beta. Both chains include structurally invariant and variable regions that participate in the binding of conventional peptide antigens. The enterotoxins are thought to link up with the beta-chain variable--or V-beta (V.sub..beta.)--region, at a part not involved in the binding of typical antigens.
Each enterotoxin interacts with particular V.sub..beta. types. For instance, one enterotoxin may be recognized by the variable types numbered 5 and 12, whereas another might be recognized by types 12, 15, and 18. For example, SEB has been shown to be specific for T cells bearing V.sub..beta. elements such as 7 and 8.1-8.3 in mice (Herman, A., J. W. Kappler, P. Marrack, A. M. Pullen [1991] Ann. Rev. Immunol. 9:745-772). Investigators estimate that every human has fewer than 30 V.sub..beta. types, although the fraction of helper T cells carrying any given type can differ from person to person. A conventional antigen can activate only the relatively few helper cells specific for that antigen. A given enterotoxin, however, can activate many times that number of helpers (having a huge variety of peptide-antigen specificities) as long as the T cells bear selected V.sub..beta. types.
Although superantigens are suspected of, at times, causing over-activation of the immune system, some evidence suggests that superantigens may also depress the immune system. T cell clones aroused by superantigens often disappear (depletion) or become inactive (anergy) after being stimulated. Staphylococcal enterotoxins, the prototype superantigens, activate T cells bearing specific T cell antigen receptor .beta.-chain variable region elements. Their V.sub..beta. specificity has profound implications with regard to expansion, anergy, and deletion of various T cell populations in terms of immunologic disease. It has been demonstrated that although an initial mitogenic effect is observed after in vivo administration of staphylococcal enterotoxin B (SEB), the lasting result appears to be both clonal anergy and deletion of V.sub..beta. specific peripheral T cells (Kawake, Y., A. Ochi [1991] Nature 349:245-248; Kawake, Y., A. Ochi [1990] J. Exp. Med. 172:1065-1070; Rellahan, B. L., L. A. Jones, A. M. Kruisbeek, A. M. Fry, L. A. Matis [1990] J. Exp. Med. 172:1092-1100).
Thus far, we have primarily focused on the interaction between superantigens and helper T cell activity; however, the possible deranging effects of superantigens on B cells should not be ignored. Staphylococcal enterotoxins sometimes enhance antibody production by B cells and sometimes inhibit it, depending on the initial state of immune arousal. Enhancement and suppression may each be destructive. Inhibition of antibody production can depress immune functioning. Overzealous production can lead to immune complex disorders, in which antibodies attract various components of the immune system to healthy tissue, clogging them and impeding normal function.
Interaction of the staphylococcal enterotoxins with class II molecules induces production of the cytokines tumor necrosis factor alpha (TNF.alpha.) and interleukin-1 (IL-1) by monocytes (Fischer, H., M. Dohlsten, U. Andersson, G. Hedlund, P. Ericsson, J. Hansson, H. O. Sjogren [1990] J. Immunol. 144:4663; Gjorloff, A., H. Fischer, G. Hedlund, J. Hansson, J. S. Kenney, A. C. Allison, H. O. Sjogren, M. Dohlsten [1991] Cell Immunol. 137:61). Both SEA and the related toxic shock syndrome toxin one (TSST-1) are potent inducers of TNF.alpha. and IL-1. Binding of these superantigens to MHC transduces a signal through the monocyte membrane which leads to tyrosine kinase activation and phosphorylation of multiple cytoplasmic proteins and monokine gene induction (Scholl, P. R., N. Trede, T. A. Chatila, R. S. Geha [1992] J. Immunol. 148:2237). Subsequently, monokines can have effects on T cells; for example, TNF.alpha. can further enhance human T cell proliferation (Yokota, S., T. D. Geppert, P. E. Lipsky [1988] J. Immunol. 140:531). IL-1 is an additional stimulator by increasing IL-2 secretion and IL-2 receptor expression. Both IL-1 and TNF.alpha. secretion may require the presence of T cells, particularly CD4.sup.+ 45RO.sup.+ memory T cells (Fischer et al., supra; Gjorloff et al., supra). A variety of peptide sequences of the superantigen SEA that participate in binding to the class II MHC molecules have previously been studied (Pontzer, C. H., J. K. Russell, H. M. Johnson [1989] I Immunol. 143:280; Pontzer, C., J. K. Russell, M. A. Jarpe, H. M. Johnson [1990] Int. Arch. Allergy Appl. Immunol. 93:107; Griggs, N. D., C. H. Pontzer, M. A. Jarpe, H. M. Johnson [1992] J. Immunol. 148:2516; Grossman, D., R. G. Cook, J. T. Sparrow, J. A. Mollick, R. R. Rich [1990] I Exp. Med. 172:1831; Grossman, D., M. Van, J. A. Mollick, S. K. Highlander, R. R. Rich [1991] J. Immunol. 147:3274).
Superantigens have been hypothesized to be associated with a number of pathological conditions. For example, superantigen alteration of the T cell repertoire has import for immunodeficiency and autoimmunity. T cells beating certain V.sub..beta. types have been implicated in various autoimmune conditions, including arthritis, lupus, and multiple sclerosis. It is conceivable, but not yet established, that over-activation of T cells by superantigens could play a role in certain autoimmune disorders.
Involvement of a predominant V.sub..beta. specific T cell population has been suggested for certain animal models of autoimmune disease. For example, experimental allergic encephalomyelitis (EAE) is an animal model for multiple sclerosis. Multiple sclerosis (MS) is a chronic, often disabling disease that attacks the central nervous system, damaging the protective coating that surrounds nerve fibers. EAE is mediated by V.sub..beta. 8.2.sup.+, CD4.sup.+ T cells in PL/J mice after injection with myelin basic protein (MBP). This limited heterogeneity of TCR usage has implicated the involvement of V.sub..beta. 8.2.sup.+, CD4.sup.+ T cells in EAE in PL/J mice immunized with rat myelin basic protein (Acha-Orbea, H., D. J. Mitchell, L. Timmerman, D. C. Wraith, G. S. Tausch, M. K. Waldon, S. S. Zamvil, H. O. McDevitt, L. Steinman [1988] Cell 54:263-273).
Recently, several novel immunological approaches have been explored relevant to autoimmune diseases such as EAE in mice and rats and lupus nephritis in MRL/lpr mice. Many have been directed toward blocking the function of the effector CD4.sup.+ T cell which has been shown to exhibit V.sub..beta. isotype restriction in EAE. These approaches have included the use of anti-TCR antibodies (Acha-Orbea et al., supra), synthetic TCR peptides (Offner, H., G. A. Hashim, .A. A. Vandenbark [1991] Science 251:430-432) and superantigen treatment (Kim, C., K. A. Siminovitch, A. Ochi [1991]J. Exp. Med. 174:1431-1437).
Superantigens are also associated with retroviruses such as mouse mammary tumor virus (MMTV), and possibly human immunodeficiency virus (HIV), the virus responsible for AIDS. It has recently been reported that two exogenous strains of MMTV encode retroviral superantigens in the open reading frames (ORFs) of the 3' Long Terminal Repeat (LTR) of the viral genome (Pullen, A. M., Y. Choi, E. Kushnir, J. Kappler, P. Marrack [1992] J. Exp. Med. 175:41-47; Choi, Y., P. Marrack, J. Kappler [1992] J. Exp. Med. 175:847-851). There is preliminary evidence that the HIV genome may also encode a superantigen. It has been suggested that an HIV superantigen may target a sub-population of CD4.sup.+ T cells for HIV viral replication (Laurence, J., A. S. Hodtsev, D. N. Posnett [1992] Nature 358:255-259). HIV infection also results in the programmed cell death of CD4.sup.+ T cells (apoptosis), both in vitro and in vivo, possibly as a result of an HIV protein with superantigen properties (Gougeon, M-L., L. Montagnier [1993] Science 260:1269-1270). Feline immunodeficiency virus (FIV) is a lentivirus which has been described extensively in the literature. See, for example, Kiyomasu, Takahiro, et al. (1991) "Identification of Feline Immunodeficiency Virus rev Gene Activity" Journal of Virology 65(8):4539-4542, and references cited therein. There has also been speculation that the human spumaretrovirus (HSRV) expresses a superantigen ("Molecular Biology of the Human Spumavirus," in Human Retroviruses, B. R. Cullen, ed., Oxford University Press, Oxford and New York, 1993, pp. 205-206).
Like many viruses, including MMTV and FIV, the HIV genome has a 3' Long Terminal Repeat. Initial studies indicated that a protein encoded by an ORF in the 3' LTR had negative effects on HIV replication in vitro, and hence was designated Negative Factor (NeF). NeF is a 25-29 kD protein that is mainly located the cytoplasm of HIV-infected cells, but is also associated with the plasma membrane (Allan, J. S., J. E. Coligan, T-H. Lee, M. F. McLane, P. J. Kanki, J. E. Groopman, M. Essex [1985] Science 230:810-813). The role of NeF in retrovital pathogenesis has been studied (Laurent, A. G., A. G. Hovanessian, Y. Riviere, b. Krust, A. Regnault, L. Montagnier, A. Findeli, M. P. Kieny, B. Guy [1990] J. Gen. Virol. 71:2273-2281). There are no reports that suggest NeF as a superantigen.